Apoptosis, or programmed cell death, is the process by which a cell will actively self-destruct in response to certain developmental stimuli. The apoptotic death process is associated with profound but well defined morphological changes in the cell, including intranucleosomal DNA fragmentation. However, apoptosis is distinguishable from necrosis, which is cell death that occurs in response to physical injury.
Apoptosis is a fundamentally important process in the regulation of cell proliferation required for normal cell and organ development (24) and regulation of apoptosis in required to maintain the ongoing stability of biological systems. Apoptosis also modulates viral pathogenesis and latency (10,16, 35). In addition, apoptosis has been strongly suggested to serve as a natural defense against cancer (21). One reason for this belief is that the molecular events that result in apoptosis are intimately related to the molecular events that lead to the development of cancer ("transformation"). Another reason is that cancerous states are often manifested by unrestricted cell division, and by the failure of cells to undergo apoptosis.
Although a number of genes that either induce or suppress apoptosis have been identified, regulation of apoptosis and its connection to transformation is poorly understood. It is known that expression of genes that appear to deregulate cell growth, such as E1A (1, 25) and c-myc (7, 26), can initiate the apoptotic response. It is also known that negative growth signals such as growth factor withdrawal (24) or induction of p53 levels (17) are similarly associated with apoptosis. These findings suggest that apoptosis may arise because of incompatible or conflicting growth signals (4, 27). These conflicting signals may arise during the transformation process.
It is also known that two putative oncogenes, bcl-2 (12), the adenovirus E1B 19K gene (1, 4, 25), can inhibit apoptosis. In transformation assays, E1B will cooperate with E1A (33), and bcl-2 will cooperate with both E1A (25) and c-myc (2, 8, 31). By such cooperation, the cellular stimulation that generally results in apoptosis is induced, but apoptosis is inhibited; these effects appear to result in the hyperactive cellular metabolism and excessive cell division that are characteristic of the cancerous state.
The nuclear phosphoprotein known as p53 has a number of recognized functions. First, p53 has a specific DNA binding capability that can both positively and negatively regulate transcription (reviewed in 29). Second, p53 is recognized as a tumor suppressor gene. Evidence for p53's role in suppressing cancer include the finding that mutations in p53 are the most prevalent genetic alterations in found in human tumors (13, 32), and the finding that the loss of p53 function greatly accelerates the frequency of tumor formation in humans (19) and animal models (6). Third, there is evidence that p53 regulates control of the cell cycle (5, 20). Fourth, p53 may be involved in apoptosis in some (but not all) cells (28). These last two functions may be related. For example, in response to DNA damage, p53 levels are increased, and p53 acts as either a cell cycle check point, producing G1 arrest to permit repair (15), or as a trigger for apoptosis (3, 18).
Prior to the discoveries upon which the present invention is based, how the apoptosis-inducing effects of E1A were mediated, or how the apoptosis-inhibiting effects of putative oncogenes E1B(19K), bcl-2, and ras were mediated, were not known.